Laurence Delhaes

The Airway Microbiota in Cystic Fibrosis: A Complex Fungal and Bacterial Community—Implications for Therapeutic Management

Background Given the polymicrobial nature of pulmonary infections in patients with cystic fibrosis (CF), it is essential to enhance our knowledge on the composition of the microbial community to improve patient management. In this study, we developed a pyrosequencing approach to extensively explore the diversity and dynamics of fungal and prokaryotic populations in CF lower airways. Methodology and Principal Findings Fungi and bacteria diversity in eight sputum samples collected from four adult CF patients was investigated using conventional microbiological culturing and high-throughput pyrosequencing approach targeting the ITS2 locus and the 16S rDNA gene. The unveiled microbial community structure was compared to the clinical profile of the CF patients. Pyrosequencing confirmed recently reported bacterial diversity and observed complex fungal communities, in which more than 60% of the species or genera were not detected by cultures. Strikingly, the diversity and species richness of fungal and bacterial communities was significantly lower in patients with decreased lung function and poor clinical status. Values of Chao1 richness estimator were statistically correlated with values of the Shwachman-Kulczycki score, body mass index, forced vital capacity, and forced expiratory volume in 1 s (p = 0.046, 0.047, 0.004, and 0.001, respectively for fungal Chao1 indices, and p = 0.010, 0.047, 0.002, and 0.0003, respectively for bacterial Chao1 values). Phylogenetic analysis showed high molecular diversities at the sub-species level for the main fungal and bacterial taxa identified in the present study. Anaerobes were isolated with Pseudomonas aeruginosa, which was more likely to be observed in association with Candida albicans than with Aspergillus fumigatus. Conclusions In light of the recent concept of CF lung microbiota, we viewed the microbial community as a unique pathogenic entity. We thus interpreted our results to highlight the potential interactions between microorganisms and the role of fungi in the context of improving survival in CF.

Occurrence and relevance of filamentous fungi in respiratory secretions of patients with cystic fibrosis — a review

The colonization of airways by filamentous fungi and the development of respiratory infections require some predisposing factors as encountered in patients with cystic fibrosis (CF). Indeed, the defective mucociliary clearance which characterizes the disease is associated with local immunological disorders. In addition, the prolonged therapy with antibiotics and the use of corticosteroid treatments also facilitate fungal growth. An important fungal biota has been described in respiratory secretions of patients suffering from CF. Aspergillus fumigatus, Scedosporium apiospermum and Aspergillus terreus for filamentous fungi and Candida albicans for yeasts are the main fungal species associated with CF. Although less common, several fungal species including Aspergillus flavus and Aspergillus nidulans may be isolated transiently from CF respiratory secretions, while others such as Exophiala dermatitidis and Scedosporium prolificans may chronically colonize the airways. Moreover, some of them like Penicillium emersonii and Acrophialophora fusispora are encountered in humans almost exclusively in the context of CF. As fungal complications in CF patients are essentially caused by filamentous fungi the present review will not include works related to yeasts. In CF patients, fungi may sometimes be responsible for deterioration of lung function, as occurs in allergic broncho-pulmonary aspergillosis (ABPA) which is the most common fungal disease in this context. Additionally, although the clinical relevance of the fungal airway colonization is still a matter of debate, filamentous fungi may contribute to the local inflammatory response, and therefore to the progressive deterioration of the lung function.

Fungal allergy in asthma–state of the art and research needs

Sensitization to fungi and long term or uncontrolled fungal infection are associated with poor control of asthma, the likelihood of more severe disease and complications such as bronchiectasis and chronic pulmonary aspergillosis. Modelling suggests that >6.5 million people have severe asthma with fungal sensitizations (SAFS), up to 50% of adult asthmatics attending secondary care have fungal sensitization, and an estimated 4.8 million adults have allergic bronchopulmonary aspergillosis (ABPA). There is much uncertainty about which fungi and fungal allergens are relevant to asthma, the natural history of sensitisation to fungi, if there is an exposure response relationship for fungal allergy, and the pathogenesis and frequency of exacerbations and complications. Genetic associations have been described but only weakly linked to phenotypes. The evidence base for most management strategies in ABPA, SAFS and related conditions is weak. Yet straightforward clinical practice guidelines for management are required. The role of environmental monitoring and optimal means of controlling disease to prevent disability and complications are not yet clear. In this paper we set out the key evidence supporting the role of fungal exposure, sensitisation and infection in asthmatics, what is understood about pathogenesis and natural history and identify the numerous areas for research studies.

Synthesis and antimalarial activity in vitro of potential metabolites of ferrochloroquine and related compounds.

In man, the two major metabolites of the antimalarial drug chloroquine (CQ) are monodesethylchloroquine (DECQ) and didesethylchloroquine (di-DECQ). By analogy with CQ, the synthesis and the in vitro tests of some amino derivatives of ferrochloroquine (FQ), a ferrocenic analogue of CQ which are presumed to be the oxidative metabolites of FQ, are reported. Desmethylferrochloroquine 1a and didesmethylferrochloroquine 2 would be more potent against schizontocides than CQ in vitro against two strains (HB3 and Dd2) of Plasmodium falciparum. Other secondary amino derivatives have been prepared and proved to be active as antimalarial agents in vitro, too.

Microplanktonic Community Structure in a Coastal System Relative to a Phaeocystis Bloom Inferred from Morphological and Tag Pyrosequencing Methods

Background Massive phytoplankton blooms, like the recurrent Phaeocystis proliferation observed every year in the Eastern English Channel (EEC), have a significant influence on the overall planktonic community structure and their food web dynamics. As well as being an important area for local fisheries, the EEC is an ideal ecosystem for work on microbial diversity. This is because, although its environmental context is relatively complex, it is reasonably well understood due to several years of monitoring and morphological observations of its planktonic organisms. The objective of our study was to better understand the under-explored microbial eukaryotic diversity relative to the Phaeocystis bloom. Methodology and Principal Findings The community structure of microplankton (diatoms, haptophytes, ciliates and dinoflagellates) was studied through morphological observations and tag pyrosequencing. During the annual Phaeocystis spring bloom, the phytoplankton biomass increased by 34-fold, while the microzooplankton biomass showed a 4-fold increase, representing on average about 4.6% of the biomass of their phytoplankton prey. Tag pyrosequencing unveiled an extensive diversity of Gymnodiniaceae, with G. spirale and G. fusiformis representing the most abundant reads. An extended diversity of Phaeocystales, with partial 18S rDNA genes sequence identity as low as 85% was found, with taxa corresponding to P. globosa, but also to unknown Phaeocystaceae. Conclusions Morphological analyses and pyrosequencing were generally in accordance with capturing frequency shifts of abundant taxa. Tag pyrosequencing allowed highlighting the maintenance of microplankton diversity during the Phaeocystis bloom and the increase of the taxa presenting low number of reads (minor taxa) along with the dominant ones in response to biotic and/or abiotic changing conditions. Although molecular approaches have enhanced our perception on diversity, it has come to light that the challenge of modelling and predicting ecological change requires the use of different complementary approaches, to link taxonomic data with the functional roles of microbes in biogeochemical cycles.

Molecular epidemiology of human Blastocystis isolates in France

Blastocystis sp. is the most common eukaryotic parasite in the intestinal tract of humans. Due to its strong impact in public health, in this study, we determined the frequency of different Blastocystis subtypes in patients in France. We hypothesized on the mode of transmission and tested a possible relationship between the subtype and symptomatic status. We obtained a total of 40 stool samples identified as positive for Blastocystis by microscopic examination of smears. Participants consisted of 25 symptomatic and 15 asymptomatic patients, for whom clinical and parasitological data were collected. For nested-polymerase chain reaction and genotyping, DNA was extracted directly from fecal samples or from fecal cultures. Morphological forms observed in fecal cultures were uncorrelated with symptomatic status. Genotyping using partial small subunit rRNA gene analysis identified a total of 43 Blastocystis isolates corresponding to 37 single infections and three mixed infections by two different subtypes. These 43 isolates belonged to five subtypes (1, 2, 3, 4, and 7) with predominance of subtype 3 (53.5%). Patient symptomatic status was uncorrelated with Blastocystis subtype.

Population-based surveillance for scedosporiosis in Australia: epidemiology, disease manifestations and emergence of Scedosporium aurantiacum infection.

Australia-wide population-based surveillance for scedosporiosis identified 180 cases, with 118 (65.6%) cases of colonization and 62 (34.4%) cases of infection. Predisposing factors for isolation of Scedosporium spp. included chronic lung disease in 37.8% and malignancy in 21.7% of cases. Predictors of invasive disease (n=62) included haematological stem cell transplantation (n=7), leukaemia (n=16) and diabetes mellitus (n=8). Of 183 phenotypically-speciated isolates, 75 (41%) were Scedosporium prolificans (risk factors: haematologic cancer (n=17), neutropaenia (n=14)) and 108 (59%) had Scedosporium apiospermum/Pseudallescheria boydii phenotype [risk factor: diabetes (n=15)]. Scedosporium prolificans (p 0.01) and leukaemia (p 0.03) independently predicted death. Epidemiological and antifungal susceptibility profiles of Scedosporium aurantiacum (prevalence>or=15.8%) and S. apiospermum were similar. No patient with S. aurantiacum infection (n=6) died. This is the first description of clinical features associated with S. aurantiacum.

Novel metallocenic compounds as antimalarial agents. Study of the position of ferrocene in chloroquine

Abstract The synthesis, characterization and antimalarial activity of two new ferrocene–chloroquine compounds are reported. One of them, 7-chloro-4- N -[(4- N′ -ethyl- N′ -ferrocenylmethyl)ammonio-1-methylbutylamino]quinolin-1-ium bi-tartrate ( 2 ) showed very promising antimalarial activity in vivo on mice infected with Plasmodium berghei N. and Plasmodium yoelii NS. and in vitro against chloroquine resistant-strains of Plasmodium falciparum .

Severe congenital toxoplasmosis due to a Toxoplasma gondii strain with an atypical genotype: case report and review.

Laurence Delhaes1,2*, Daniel Ajzenberg2,3,4, Bérangère Sicot5, Philippe Bourgeot5, Marie-Laure Dardé2,3,4, Eduardo Dei-Cas1,2 and Véronique Houfflin-Debarge5 1Parasitology-Mycology Service (EA3609), Faculty of Medicine, University of Nord de France (UDSL), University Hospital Centre & IFR-142, Institut Pasteur de Lille, Lille, France 2Centre National de Référence (CNR) Toxoplasmose/Toxoplasma Biological Resource Center (BRC), Limoges 87025, France 3Département de Parasitologie-Mycologie, Centre Hospitalier-Universitaire Dupuytren, Limoges 87042, France 4Laboratoire de Parasitologie-Mycologie, EA 3174-NETEC, Faculté de Médecine, Université de Limoges, Limoges 87025, France 5Department of Obstetrics, Lille Hospital, Lille University, Lille, France

The lung mycobiome: an emerging field of the human respiratory microbiome

The lung microbiome, which is believed to be stable or at least transient in healthy people, is now considered as a poly-microorganism component contributing to disease pathogenesis. Most research studies on the respiratory microbiome have focused on bacteria and their impact on lung health, but there is evidence that other non-bacterial organisms, comprising the viruses (virome) and fungi (mycobiome), are also likely to play an important role in healthy people as well as in patients. In the last few years, the lung mycobiome (previously named the fungal microbiota or microbiome) has drawn closer attention. There is growing evidence that the lung mycobiome has a significant impact on clinical outcome of chronic respiratory diseases (CRD) such as asthma, chronic obstructive pulmonary disease, cystic fibrosis, and bronchiectasis. Thanks to advances in culture independent methods, especially next generation sequencing, a number of fungi not detected by culture methods have been molecularly identified in human lungs. It has been shown that the structure and diversity of the lung mycobiome vary in different populations (healthy and different diseased individuals) which could play a role in CRD. Moreover, the link between lung mycobiome and different biomes of other body sites, especially the gut, has also been unraveled. By interacting with the bacteriome and/or virome, the respiratory mycobiome appears to be a cofactor in inflammation and in the host immune response, and therefore may contribute to the decline of the lung function and the disease progression. In this review, we report the recent limited explorations of the human respiratory mycobiome, and discuss the mycobiome’s connections with other local microbial communities, as well as the relationships with the different biomes of other body sites. These studies suggest several outlooks for this understudied emerging field, which will certainly call for a renewal of our understanding of pulmonary diseases.